Crystallographic Structure of Xanthorhodopsin, the Light-Driven Proton Pump with a Dual Chromophore

Total Page:16

File Type:pdf, Size:1020Kb

Crystallographic Structure of Xanthorhodopsin, the Light-Driven Proton Pump with a Dual Chromophore Crystallographic structure of xanthorhodopsin, the light-driven proton pump with a dual chromophore Hartmut Lueckea,b,c,1, Brigitte Schobertb, Jason Stagnoa, Eleonora S. Imashevab, Jennifer M. Wangb, Sergei P. Balashovb,1, and Janos K. Lanyib,c,1 Departments of aMolecular Biology and Biochemistry, and bPhysiology and Biophysics, and cCenter for Biomembrane Systems, University of California, Irvine, CA 92697 Edited by Harry B. Gray, California Institute of Technology, Pasadena, CA, and approved September 16, 2008 (received for review July 23, 2008) Homologous to bacteriorhodopsin and even more to proteorho- found recently in the genome of an abundant coastal ocean dopsin, xanthorhodopsin is a light-driven proton pump that, in methylotroph (11) and earlier in the genomes of Gloebacter addition to retinal, contains a noncovalently bound carotenoid violaceous, Pyrocystis lunula (12), and others. The proteins in this with a function of a light-harvesting antenna. We determined the clade exhibit significantly less homology to the proteorhodopsins structure of this eubacterial membrane protein–carotenoid com- (11). For example, 137 residues (50%) are identical in plex by X-ray diffraction, to 1.9-Å resolution. Although it contains Gloeobacter rhodopsin and xanthorhodopsin, but only 60 resi- 7 transmembrane helices like bacteriorhodopsin and archaerho- dues (22%) in proteorhodopsin and xanthorhodopsin. Although dopsin, the structure of xanthorhodopsin is considerably different considerable sequence differences separate xanthorhodopsin from the 2 archaeal proteins. The crystallographic model for this from the proteorhodopsins (Fig. 1), its structure, the first for a rhodopsin introduces structural motifs for proton transfer during eubacterial proton pump, is likely to be relevant to other the reaction cycle, particularly for proton release, that are dramat- eubacterial retinal-based pumps. ically different from those in other retinal-based transmembrane pumps. Further, it contains a histidine–aspartate complex for Results and Discussion regulating the pKa of the primary proton acceptor not present in Xanthorhodopsin was crystallized from bicelles (13), with a type archaeal pumps but apparently conserved in eubacterial pumps. In I arrangement of stacked bilayers. The structure was solved to addition to aiding elucidation of a more general proton transfer 1.9-Å resolution (Table 1). The P1 unit cell contains 2 molecules mechanism for light-driven energy transducers, the structure de- of xanthorhodopsin with a head-to-tail arrangement somewhat fines also the geometry of the carotenoid and the retinal. The close similar to 2-dimensional crystal forms of bacteriorhodopsin (14) approach of the 2 polyenes at their ring ends explains why the and halorhodopsin (15), as well as 3-dimensional crystals of the efficiency of the excited-state energy transfer is as high as Ϸ45%, D85S bacteriorhodopsin mutant (16), sensory rhodopsin II (17, and the 46° angle between them suggests that the chromophore 18), and Anabaena sensory rhodopsin (19). Considering its location is a compromise between optimal capture of light of all function as an ion transporter in the cell membrane, xanthor- polarization angles and excited-state energy transfer. hodopsin is unlikely to form such dimers in the original Salini- bacter cells. carotenoid antenna ͉ energy transfer ͉ retinal protein ͉ salinixanthin ͉ X-ray structure Location and Binding Site of Carotenoid Antenna. The C40 carot- enoid lies transverse against the outer surface of helix F at a Ϸ54° ncreased efficiency of light harvesting by antennae, such as angle to the membrane normal, buried at the protein–lipid carotenoids, is common in photosynthetic membranes, large boundary (Fig. 2A). Its keto-ring is immobilized by residues at I ␤ multiprotein systems that contain tens or hundreds of chro- the extracellular ends of helices E and F and by the -ionone ring mophores. The much simpler retinal-based archaeal proton of the retinal (Fig. 2B) and rotated 82° from the plane of the methyl side chains of its polyene chain and therefore from the pumps bacteriorhodopsin and archaerhodopsin lack antennae; ␲ A light collection and proton translocation are performed by a plane of the extended -system (Fig. 2C). The ring C O is not single protein with a single chromophore. Recently, it was shown hydrogen-bonded. The immobilized and acutely out-of-plane orientation of the keto-ring minimizes participation of its 2 that a light-harvesting antenna can function also in a retinal ␲ protein. Xanthorhodopsin (1), of the eubacterium Salinibacter double bonds in the conjugated -system and explains the ruber (2), contains a single energy-donor carotenoid, salinixan- well-resolved vibronic bands of the carotenoid, the lack of a red-shift of the absorption bands upon binding, and the strong thin (3), and a single acceptor, retinal, in a small (25 kDa) BIOPHYSICS membrane protein. Because energy transfer is from the short- CD bands in the visible region (5). The relatively rigid polyene is wedged in a slot on the outside of helix F, with one side formed lived S2 carotenoid level (4), there must be a short distance and favorable geometry between the 2 chromophores to account for by the Leu-194 and Leu-197 side chains and the other by the Ile-205 side chain, and its base by Gly-201. The carotenoid its high (40–50%) efficiency. Close interaction of the 2 chro- A mophores is indicated by dependence of the carotenoid confor- glucoside is hydrogen-bonded to the C O and the NH2 of the mation on the presence of the retinal in the protein (1, 5, 6) and amide side chain of Asn-191, as well as NH1 of Arg-184. The spectral changes of the carotenoid during the photochemical transformations of the retinal (1), but, as for the proteorhodop- Author contributions: S.P.B. and J.K.L. designed research; B.S., E.S.I., J.M.W., S.P.B., and sin family of proteins, no direct structural information has been J.K.L. performed research; E.S.I., J.M.W., and S.P.B. contributed new reagents/analytic tools; available (4, 7). Unexpectedly, the crystallographic structure of H.L., J.S., and J.K.L. analyzed data; and H.L., S.P.B., and J.K.L. wrote the paper. xanthorhodopsin we report here reveals not only the location of The authors declare no conflict of interest. the antenna but also striking differences from the archaeal This article is a PNAS Direct Submission. retinal proteins, bacteriorhodopsin and archaerhodopsin. Data deposition: The atomic coordinates have been deposited in the Protein Data Bank, The photocycle of xanthorhodopsin (8) and the functional www.pdb.org (PDB ID code 3DDL). residues in the ion transfer pathway (1) are similar to those of the 1To whom correspondence may be addressed. E-mail: [email protected], [email protected], or numerous other eubacterial proton pumps, the proteorho- [email protected]. dopsins (9, 10). Proteins homologous to xanthorhodopsin were © 2008 by The National Academy of Sciences of the USA www.pnas.org͞cgi͞doi͞10.1073͞pnas.0807162105 PNAS ͉ October 28, 2008 ͉ vol. 105 ͉ no. 43 ͉ 16561–16565 Downloaded by guest on September 29, 2021 Fig. 1. Sequence alignment of green light-absorbing proteorhodopsin (PR), xanthorhodopsin (XR), and bacteriorhodopsin (BR), reevaluated from the one shown in ref. 1 by using information gained from the diffraction structure. Red, conserved residues in all three; purple, conserved residues in xanthorho- dopsin and bacteriorhodopsin; yellow, conserved residues in xanthorhodop- sin and proteorhodopsins; blue, residues involved with carotenoid binding. Top row of numbers refer to the xanthorhodopsin sequence; bottom row to the bacteriorhodopsin sequence. Underlining indicates residues in transmem- brane helices. Proteorhodopsin sequence refers to a species from Monterey Bay, MBP1 (protein accession No. AAG10475). dependence of the carotenoid spectrum on the retinal is ex- plained by the fact that the retinal ␤-ionone ring is part of the carotenoid binding site (Fig. 2B). The keto-ring of the carotenoid is in the space occupied by Trp-138 in bacteriorhodopsin, one of the bulky side chains that confines the retinal ␤-ionone ring in that protein. In xanthor- Table 1. Data collection and refinement statistics Data collection Beamline 9.1, SSRL, Menlo Park, CA Wavelength, Å 0.979 Fig. 2. Location of salinixanthin (orange) and retinal (magenta) in xanthor- Space group P1 hodopsin. (A) The extended carotenoid is tightly bound on the transmem- Cell dimensions a ϭ 52.7 Å, b ϭ 59.5 Å, c ϭ 59.7 Å brane surface of xanthorhodopsin, traversing nearly the entire bilayer, with ␣ ϭ 76.4°, ␤ ϭ 74.9°, ␥ ϭ 64.1° an inclination of 54° to the membrane normal. Its keto-ring binds in a pocket Resolution range, Å 45.10–1.90 between helices E and F, very near the ␤-ionone ring of the retinal. The angle Total reflections 166,560 between the chromophore axes is 46°. The angle between the planes of their Unique reflections 46,289 ␲-systems is 68°. Horizontal lines indicate the approximate boundaries of the Redundancy 3.6 (3.5)* lipid bilayer. Helices E and F are marked. (B) The binding pocket of the keto Completeness, % 94.1 (85.5)* ring is formed by Leu-148, Gly-156, Phe-157, Thr-160, Met-208, and Met-211, ␤ Mean I/␴ 8.4 (1.5)* as well as the retinal -ionone ring. (C) The keto-ring of the carotene is rotated 82° out of plane of the salinixanthin-conjugated system and is in van der Waals Rsym,% 5.7 (48.5)* distance of the retinal ␤-ionone and the phenolic side chain of Tyr-207. Refinement Resolution range, Å 45.10–1.90 No. of reflections used 46,278 hodopsin, it is replaced by a glycine. This residue replacement R /R ,%† 24.7/26.5 work free might be the best diagnostic in a sequence for the possibility of Rmsd bonds, Å 0.012 Rmsd angles, ° 1.35 binding a salinixanthin-like carotenoid.
Recommended publications
  • POSTER LIST (Alphabetical by Author)
    POSTER LIST (alphabetical by author) First author Poster Title Session or General Archibald, Kevin Modeling the impact of zooplankton diel vertical migration on General the carbon export flux of the biological pump Arroyo, Mar CO2 System Dynamics in the Dalton Polynya, East Antarctica Polar Bif, Mariana Using cruises and Bio-Argo floats data to estimate dissolved Time-series organic carbon in the Northeast Pacific Ocean Bochdansky, Net carbon remineralization (defined here as CO2 release) of Microzooplankton Alexander stable isotope-labeled phytoplankton measured in the mesopelagic environment with a newly designed deep-sea incubator Bociu, Ioana Initial DECK analysis of the carbon cycle in GFDL's CM4 Time-series contribution to CMIP6 Boyd, Philip A web- based Best Practice Guide to design marine multiple General driver experiments Buchanan, Marine nitrogen fixers crucial for dust-driven strengthening of Phytoplankton/models Pearse the biological carbon pump Busecke, Julius How important are forced changes in the equatorial current General system for the extent of tropical oxygen minimum zones? Cael, B.B. A metabolic model of biogeochemistry Phytoplankton/models Campbell, Mesozooplankton are not herbivores: The importance of Microzooplankton Robert microzooplankton in mesozooplankton diets and in arctic and *C. Ashjian, sub-arctic trophic linkages presenting Chappell, P. Both dissolved iron and coastal water additions to Southern Polar Dreux Drake Passage waters stimulate similar growth responses and shifts in diatom community composition.
    [Show full text]
  • A Voltage-Dependent Fluorescent Indicator for Optogenetic Applications, Archaerhodopsin-3: Structure and Optical Properties
    F1000Research 2017, 6:33 Last updated: 30 JUL 2021 RESEARCH NOTE A voltage-dependent fluorescent indicator for optogenetic applications, archaerhodopsin-3: Structure and optical properties from in silico modeling [version 3; peer review: 3 approved] Dmitrii M. Nikolaev1, Anton Emelyanov1, Vitaly M. Boitsov1, Maxim S Panov2, Mikhail N. Ryazantsev 2,3 1Saint-Petersburg National Research Academic University of the Russian Academy of Science, St. Petersburg, Russian Federation 2Saint-Petersburg State University, St. Petersburg, Russian Federation 3Saint-Petersburg Scientific Center of the Russian Academy of Sciences, St. Petersburg, Russian Federation v3 First published: 11 Jan 2017, 6:33 Open Peer Review https://doi.org/10.12688/f1000research.10541.1 Second version: 17 Jan 2017, 6:33 https://doi.org/10.12688/f1000research.10541.2 Reviewer Status Latest published: 15 Nov 2017, 6:33 https://doi.org/10.12688/f1000research.10541.3 Invited Reviewers 1 2 3 Abstract It was demonstrated in recent studies that some rhodopsins can be version 3 used in optogenetics as fluorescent indicators of membrane voltage. (revision) report report One of the promising candidates for these applications is 15 Nov 2017 archaerhodopsin-3. While it has already shown encouraging results, there is still a large room for improvement. One of possible directions version 2 is increasing the intensity of the protein's fluorescent signal. Rational (revision) report report report design of mutants with an improved signal is an important task, which 17 Jan 2017 requires both experimental and theoretical studies. Herein, we used a homology-based computational approach to predict the three- version 1 dimensional structure of archaerhodopsin-3, and a Quantum 11 Jan 2017 Mechanics/Molecular Mechanics (QM/MM) hybrid approach with high- level multireference ab initio methodology (SORCI+Q/AMBER) to 1.
    [Show full text]
  • Ecosystem Structure and Dynamics in the North Pacific Subtropical Gyre
    Ecosystems DOI: 10.1007/s10021-017-0117-0 Ó 2017 The Author(s). This article is published with open access at Springerlink.com 20th Anniversary Paper Ecosystem Structure and Dynamics in the North Pacific Subtropical Gyre: New Views of an Old Ocean David M. Karl1,2* and Matthew J. Church1,2,3 1Center for Microbial Oceanography: Research and Education, C-MORE Hale, University of Hawaii at Manoa, 1950 East-West Rd., Honolulu, Hawaii 96822, USA; 2School of Ocean and Earth Science and Technology, University of Hawaii at Manoa, Honolulu, Hawaii 96822, USA; 3Present address: Flathead Lake Biological Station, University of Montana, Polson, MT 59860, USA ABSTRACT The North Pacific Subtropical Gyre (NPSG) is one of community with a relatively stable plankton com- the largest biomes on Earth. It has a semi-enclosed munity structure and relatively low variability in key surface area of about 2 9 107 km2 and mean depth of microbiological rates and processes. Now, after nearly nearly 5 km and includes a broad range of habitats three decades of observations and experimentation from warm, light-saturated, nutrient-starved surface we present a new view of this old ocean, one that waters to the cold, nutrient-rich abyss. Microorgan- highlights temporal variability in ecosystem pro- isms are found throughout the water column and are cesses across a broad range of scales from diel to vertically stratified by their genetically determined decadal and beyond. Our revised paradigm is built on metabolic capabilities that establish physiological the strength of high-quality time-series observations, tolerances to temperature, light, pressure, as well as on insights from the application of state-of-the-art organic and inorganic growth substrates.
    [Show full text]
  • Han-Optogenetic-Review-ACS-2012.Pdf
    Review pubs.acs.org/chemneuro In Vivo Application of Optogenetics for Neural Circuit Analysis Xue Han Biomedical Engineering Department, Boston University, Boston, Massachusetts, United States ABSTRACT: Optogenetics combines optical and genetic methods to rapidly and reversibly control neural activities or other cellular functions. Using genetic methods, specific cells or anatomical pathways can be sensitized to light through exogenous expression of microbial light activated opsin proteins. Using optical methods, opsin expressing cells can be rapidly and reversibly controlled by pulses of light of specific wavelength. With the high spatial temporal precision, optogenetic tools have enabled new ways to probe the causal role of specific cells in neural computation and behavior. Here, we overview the current state of the technology, and provide a brief introduction to the practical considerations in applying optogenetics in vivo to analyze neural circuit functions. KEYWORDS: Channelrhodopsin, archaerhodopsin, halorhodopsin, cell type specificity ptogenetics is a new field being rapidly established upon algae), and others that mediate a variety of cellular functions O the first demonstration of precise activation of neurons (for reviews on opsin structure and function, see refs 9 and 10). expressing a light-activated microbial opsin, channelrhodopsin- Most sensory rhodopsins function through recruiting intra- 2, with pulses of blue light in 2005.1 Microbial (type I) opsins cellular signaling molecules without direct ion transport are classes of monolithic light activated proteins, encoded by function. However, channelrhodopsins can mediate phototaxis small genes of under a kilobase long. Three major classes of as a light-gated cation channel at high light intensity and as a microbial opsins have been adapted to optogenetically control calcium channel at low light intensity.11 cellular functions, channelrhodopsins, halorhodopsins, and Microbial opsins share sequence homology and are archaerhodopsins (Figure 1).
    [Show full text]
  • High-Performance Genetically Targetable Optical Neural Silencing by Light-Driven Proton Pumps
    Vol 463 | 7 January 2010 | doi:10.1038/nature08652 LETTERS High-performance genetically targetable optical neural silencing by light-driven proton pumps Brian Y. Chow1,2*, Xue Han1,2*, Allison S. Dobry1,2, Xiaofeng Qian1,2, Amy S. Chuong1,2, Mingjie Li1,2, Michael A. Henninger1,2, Gabriel M. Belfort2, Yingxi Lin2, Patrick E. Monahan1,2 & Edward S. Boyden1,2 The ability to silence the activity of genetically specified neurons in screen, as did two other proton pumps, the Leptosphaeria maculans a temporally precise fashion would provide the opportunity to opsin (Mac/LR/Ops)4 and cruxrhodopsin-1 (ref. 10) (albeit less than investigate the causal role of specific cell classes in neural compu- that of Arch; Fig. 1a). All light-driven chloride pumps assessed had tations, behaviours and pathologies. Here we show that members lower screen photocurrents than these light-driven proton pumps. of the class of light-driven outward proton pumps can mediate Arch is a yellow–green light-sensitive (Fig. 1b) opsin that seems to powerful, safe, multiple-colour silencing of neural activity. The express well on the neural plasma membrane (Fig. 1c; see Sup- gene archaerhodopsin-3 (Arch)1 from Halorubrum sodomense plementary Notes on Arch expression levels and enhancing Arch enables near-100% silencing of neurons in the awake brain when membrane trafficking). Arch-mediated currents exhibited excellent virally expressed in the mouse cortex and illuminated with yellow kinetics of light-activation and post-light recovery. After illumination, light. Arch mediates currents of several hundred picoamps at low Arch currents rose with a 15–85% onset time of 8.8 6 1.8 ms light powers, and supports neural silencing currents approaching (mean 6 standard error (s.e.) reported throughout, unless otherwise 900 pA at light powers easily achievable in vivo.
    [Show full text]
  • Illuminating Brain Activities with Fluorescent Protein-Based Biosensors
    chemosensors Review Illuminating Brain Activities with Fluorescent Protein-Based Biosensors Zhijie Chen 1 ID , Tan M. Truong 2 and Hui-wang Ai 2,3,4,* ID 1 California Institute for Quantitative Biosciences, QB3, University of California, Berkeley, CA 94720, USA; [email protected] 2 Center for Membrane and Cell Physiology, and Biomedical Sciences (BIMS) Graduate Program, University of Virginia, Charlottesville, VA 22908, USA; [email protected] 3 Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA 22908, USA 4 Department of Chemistry, University of Virginia, Charlottesville, VA 22908, USA * Correspondence: [email protected] Received: 9 October 2017; Accepted: 22 November 2017; Published: 28 November 2017 Abstract: Fluorescent protein-based biosensors are indispensable molecular tools for life science research. The invention and development of high-fidelity biosensors for a particular molecule or molecular event often catalyze important scientific breakthroughs. Understanding the structural and functional organization of brain activities remain a subject for which optical sensors are in desperate need and of growing interest. Here, we review genetically encoded fluorescent sensors for imaging neuronal activities with a focus on the design principles and optimizations of various sensors. New bioluminescent sensors useful for deep-tissue imaging are also discussed. By highlighting the protein engineering efforts and experimental applications of these sensors, we can consequently analyze factors influencing their performance. Finally, we remark on how future developments can fill technological gaps and lead to new discoveries. Keywords: fluorescent proteins; biosensors; fluorescent probes/indicators; brain imaging; genetically encoded calcium indicators; genetically encoded voltage sensors; genetically encoded glutamate sensors; synaptic activity indicators; pH indicators; bioluminescent sensors 1.
    [Show full text]
  • Proteorhodopsin Phototrophy in the Ocean
    letters to nature 17. Schatz, J. F. & Simmons, G. Thermal conductivity of Earth materials at high temperatures. J. Geophys. an H+-ATPase4. Similar rhodopsin-mediated, light-driven proton Res. 77, 6966±6983 (1972). pumping, formerly thought to exist only in halophilic archaea, has 18. Holt, J. B. Thermal diffusivity of olivine. Earth Planet. Sci. Lett. 27, 404±408 (1975). 1 19. Seipold, U. Temperature dependence of thermal transport properties of crystalline rocks±general law. been discovered in an uncultivated marine bacterium of the Tectonophysics 291, 161±171 (1998). `SAR86' phylogenetic group5. This ®nding suggested that a pre- 20. Hofmeister, A. M. Mantle values of thermal conductivity and the geotherm from phonon lifetimes. viously unrecognized phototrophic pathway may occur in the Science 283, 1699±1706 (1999). 21. Klemens, P. G. Thermal conductivity and lattice vibrational modes. Solid State Phys. 7, 1±98 (1958). ocean's photic zone; however, all earlier data are based solely on 22. Bouhifd, M. A., Andrault, D., Fiquet, G. & Richet, P.Thermal expansion of forsterite up to the melting recombinant DNA and in vitro biochemical analyses, and this point. Geophys. Res. Lett. 23, 134±1136 (1996). phenomenon has not yet been observed in the sea. 23. Vauchez, A., Barruol, G. & Tommasi,A. Why do continents break up parallel to ancient orogenic belts? To test whether rhodopsin-like molecules form photoactive Terra Nova 9, 62±66 (1997). 24. Tommasi, A. & Vauchez, A. Continental rifting parallel to ancient collisional belts: An effect of the proteins in native marine bacteria, we analysed membrane prepara- mechanical anisotropy of the lithospheric mantle.
    [Show full text]
  • Lokiarchaeota Archaeon Schizorhodopsin-2 (Laszr2) Is an Inward Proton Pump Displaying a Characteristic Feature of Acid-Induced S
    www.nature.com/scientificreports OPEN Lokiarchaeota archaeon schizorhodopsin‑2 (LaSzR2) is an inward proton pump displaying a characteristic feature of acid‑induced spectral blue‑shift Keiichi Kojima1,5, Susumu Yoshizawa2,5, Masumi Hasegawa2, Masaki Nakama1, Marie Kurihara1, Takashi Kikukawa3,4 & Yuki Sudo1,2* The photoreactive protein rhodopsin is widespread in microorganisms and has a variety of photobiological functions. Recently, a novel phylogenetically distinctive group named ‘schizorhodopsin (SzR)’ has been identifed as an inward proton pump. We performed functional and spectroscopic studies on an uncharacterised schizorhodopsin from the phylum Lokiarchaeota archaeon. The protein, LaSzR2, having an all‑trans-retinal chromophore, showed inward proton pump activity with an absorption maximum at 549 nm. The pH titration experiments revealed that the protonated Schif base of the retinal chromophore (Lys188, pKa = 12.3) is stabilised by the deprotonated counterion (presumably Asp184, pKa = 3.7). The fash‑photolysis experiments revealed the presence of two photointermediates, K and M. A proton was released and uptaken from bulk solution upon the formation and decay of the M intermediate. During the M‑decay, the Schif base was reprotonated by the proton from a proton donating residue (presumably Asp172). These properties were compared with other inward (SzRs and xenorhodopsins, XeRs) and outward proton pumps. Notably, LaSzR2 showed acid‑induced spectral ‘blue‑shift’ due to the protonation of the counterion, whereas outward proton pumps showed opposite shifts (red‑shifts). Thus, we can distinguish between inward and outward proton pumps by the direction of the acid‑induced spectral shift. Sunlight provides a fundamental source for organisms, including humans. Many organisms possess a variety of photoreceptor proteins that can capture sunlight.
    [Show full text]
  • Proteorhodopsin Variability and Distribution in the North Pacific
    The ISME Journal (2018) 12:1047–1060 https://doi.org/10.1038/s41396-018-0074-4 ARTICLE Proteorhodopsin variability and distribution in the North Pacific Subtropical Gyre 1 2 1 2,3 1 Daniel K. Olson ● Susumu Yoshizawa ● Dominique Boeuf ● Wataru Iwasaki ● Edward F. DeLong Received: 30 August 2017 / Revised: 21 November 2017 / Accepted: 5 December 2017 / Published online: 23 February 2018 © The Author(s) 2018. This article is published with open access Abstract Proteorhodopsin is a light-activated retinal-containing proton pump found in many marine bacteria. These photoproteins are globally distributed in the ocean’s photic zone and are capable of generating a proton motive force across the cell membrane. We investigated the phylogenetic diversity, distribution, and abundance of proteorhodopsin encoding genes in free-living bacterioplankton in the North Pacific Subtropical Gyre, leveraging a gene catalog derived from metagenomic samples from the ocean’s surface to 1000 m depth. Proteorhodopsin genes were identified at all depths sampled, but were most abundant at depths shallower than 200 m. The majority of proteorhodopsin gene sequences (60.9%) belonged to members of the SAR11 lineage, with remaining sequences distributed among other diverse taxa. We observed variations in the conserved residues fi 1234567890();,: involved in ion pumping and spectral tuning, and biochemically con rmed four different proton pumping proteorhodopsin motifs, including one unique to deep-water SAR11. We also identified a new group of putative proteorhodopsins having unknown function. Our results reveal a broad organismal and unexpected depth distribution for different proteorhodopsin types, as well as substantial within-taxon variability. These data provide a framework for exploring the ecological relevance of proteorhodopsins and their spatiotemporal variation and function in heterotrophic bacteria in the open ocean.
    [Show full text]
  • Seasonality of Archaeal Proteorhodopsin and Associated Marine Group Iib Ecotypes (Ca
    The ISME Journal (2021) 15:1302–1316 https://doi.org/10.1038/s41396-020-00851-4 ARTICLE Seasonality of archaeal proteorhodopsin and associated Marine Group IIb ecotypes (Ca. Poseidoniales) in the North Western Mediterranean Sea 1,5 1 2 3 4 Olivier Pereira ● Corentin Hochart ● Dominique Boeuf ● Jean Christophe Auguet ● Didier Debroas ● Pierre E. Galand 1 Received: 23 July 2020 / Revised: 9 November 2020 / Accepted: 18 November 2020 / Published online: 7 December 2020 © The Author(s), under exclusive licence to International Society for Microbial Ecology 2020 Abstract The Archaea Marine Group II (MGII) is widespread in the world’s ocean where it plays an important role in the carbon cycle. Despite recent discoveries on the group’s metabolisms, the ecology of this newly proposed order (Candidatus Poseidoniales) remains poorly understood. Here we used a combination of time-series metagenome-assembled genomes (MAGs) and high-frequency 16S rRNA data from the NW Mediterranean Sea to test if the taxonomic diversity within the MGIIb family (Candidatus Thalassarchaeaceae) reflects the presence of different ecotypes. The MAGs’ seasonality revealed 1234567890();,: 1234567890();,: a MGIIb family composed of different subclades that have distinct lifestyles and physiologies. The vitamin metabolisms were notably different between ecotypes with, in some, a possible link to sunlight’s energy. Diverse archaeal proteorhodopsin variants, with unusual signature in key amino acid residues, had distinct seasonal patterns corresponding to changing day length. In addition, we show that in summer, archaea, as opposed to bacteria, disappeared completely from surface waters. Our results shed light on the diversity and the distribution of the euryarchaeotal proteorhodopsin, and highlight that MGIIb is a diverse ecological group.
    [Show full text]
  • Atomistic Design of Microbial Opsin-Based Blue-Shifted Optogenetics Tools
    ARTICLE Received 4 Nov 2014 | Accepted 14 Apr 2015 | Published 15 May 2015 DOI: 10.1038/ncomms8177 OPEN Atomistic design of microbial opsin-based blue-shifted optogenetics tools Hideaki E. Kato1,*,w, Motoshi Kamiya2,*, Seiya Sugo2, Jumpei Ito3, Reiya Taniguchi1, Ayaka Orito3, Kunio Hirata4, Ayumu Inutsuka5, Akihiro Yamanaka5, Andre´s D. Maturana3, Ryuichiro Ishitani1, Yuki Sudo6, Shigehiko Hayashi2 & Osamu Nureki1 Microbial opsins with a bound chromophore function as photosensitive ion transporters and have been employed in optogenetics for the optical control of neuronal activity. Molecular engineering has been utilized to create colour variants for the functional augmentation of optogenetics tools, but was limited by the complexity of the protein–chromophore inter- actions. Here we report the development of blue-shifted colour variants by rational design at atomic resolution, achieved through accurate hybrid molecular simulations, electrophysiology and X-ray crystallography. The molecular simulation models and the crystal structure reveal the precisely designed conformational changes of the chromophore induced by combinatory mutations that shrink its p-conjugated system which, together with electrostatic tuning, produce large blue shifts of the absorption spectra by maximally 100 nm, while maintaining photosensitive ion transport activities. The design principle we elaborate is applicable to other microbial opsins, and clarifies the underlying molecular mechanism of the blue-shifted action spectra of microbial opsins recently isolated from natural sources. 1 Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan. 2 Department of Chemistry, Graduate School of Science, Kyoto University, Kyoto 606-8502, Japan. 3 Department of Bioengineering Sciences, Graduate School of Bioagricultural Sciences, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8601, Japan.
    [Show full text]
  • Bright and Fast Multicoloured Voltage Reporters Via Electrochromic FRET
    ARTICLE Received 18 Mar 2014 | Accepted 8 Jul 2014 | Published 13 Aug 2014 DOI: 10.1038/ncomms5625 Bright and fast multicoloured voltage reporters via electrochromic FRET Peng Zou1,*, Yongxin Zhao2,*, Adam D. Douglass3, Daniel R. Hochbaum1, Daan Brinks1, Christopher A. Werley1, D. Jed Harrison2, Robert E. Campbell2 & Adam E. Cohen1,4 Genetically encoded fluorescent reporters of membrane potential promise to reveal aspects of neural function not detectable by other means. We present a palette of multicoloured brightly fluorescent genetically encoded voltage indicators with sensitivities from 8–13% DF/F per 100 mV, and half-maximal response times from 4–7 ms. A fluorescent protein is fused to an archaerhodopsin-derived voltage sensor. Voltage-induced shifts in the absorption spectrum of the rhodopsin lead to voltage-dependent nonradiative quenching of the appended fluorescent protein. Through a library screen, we identify linkers and fluorescent protein combinations that report neuronal action potentials in cultured rat hippocampal neurons with a single-trial signal-to-noise ratio from 7 to 9 in a 1 kHz imaging bandwidth at modest illumination intensity. The freedom to choose a voltage indicator from an array of colours facilitates multicolour voltage imaging, as well as combination with other optical reporters and optogenetic actuators. 1 Departments of Chemistry and Chemical Biology and Physics, Harvard Stem Cell Institute, Harvard University, Cambridge, Massachusetts 02138, USA. 2 Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2G2. 3 Department of Neurobiology and Anatomy, University of Utah, Salt Lake City, Utah 84132, USA. 4 Howard Hughes Medical Institute, Harvard University, 12 Oxford Street, Cambridge, Massachusetts 02138, USA.
    [Show full text]